EP2618808A1

EP2618808A1 - Cosmetic use of dermicidin, and analogues or fragments thereof

Info

Publication number: EP2618808A1
Application number: EP11773870.8A
Authority: EP
Inventors: Caroline Delattre; Audrey Gueniche; Isabelle Castiel; Dominique Bernard
Original assignee: LOreal SA
Current assignee: LOreal SA
Priority date: 2010-09-24
Filing date: 2011-09-23
Publication date: 2013-07-31
Anticipated expiration: 2031-09-23
Also published as: US20130324476A1; WO2012038929A1; EP2618808B1; ES2605957T3

Abstract

The present invention relates, in particular, to the use of a sequence of amino acids of dermicidin or an analogue or fragment thereof, and at least one sequence of nucleic acids coding said sequence as a biomarker for ageing skin and/or the signs of skin ageing which are possibly associated with skin dryness.

Description

The present invention relates to the field of biomarkers of the skin, and more particularly of the aged skin, associated or not with a dry skin, and their implementation as targets or cosmetic active agents.

The epidermis, the superficial part of the skin, is a tissue whose cells are contiguous and integral with one another and rest on a basement membrane. It forms an outer coating comprising sebaceous or sweat glands and hair follicles.

More specifically, the epidermis is a structure whose homeostasis results from the implementation of a finely regulated set of intracellular and extracellular signals acting at all stages of proliferation, migration, cell differentiation, as well as of the synthesis of the different components of the extracellular matrix. These signals may, in particular, result from the action of factors produced by the keratinocytes.

The epidermis is conventionally divided into a basal layer of keratinocytes containing, in particular, cutaneous stem cells and constituting the germinal layer of the epidermis, a so-called spinous layer consisting of several layers of polyhedral cells arranged on the basal layer, a so-called granulosa comprising one to three layers of flattened cells containing distinct cytoplasmic inclusions, keratohyaline grains, and finally, a set of upper layers, called stratum corneum, or corneal layer, consisting of keratinocytes at the end stage of their differentiation, called corneocytes .

The stratum corneum, the outermost part of the skin that provides the barrier function between the body and the environment, and the hair shaft, the emergent part of the hair follicle that makes up the hair, are both the culmination of the healing process. differentiation of keratinocytes. Epidermal differentiation follows a maturation process in which basal layer keratinocytes differentiate and migrate to form corneocytes, dead cells that are completely keratinized. This differentiation is the result of perfectly coordinated phenomena that will lead to the maintenance of homeostasis of the epidermis, and give the skin a healthy, youthful, luminous and smooth appearance.

During aging, many physiological alterations of the skin, resulting from a dysfunction of the homeostasis of the epidermis, and in particular a dysfunction of epithelial differentiation of keratinocytes and / or proteoglycan synthesis, occur.

The alterations of epidemic homeostasis occurring during aging are mainly reflected by a decrease in the differentiation of keratinocytes, resulting in a deficit of the protein matrix of the horny cells, by an increase in the metalloproteases, and their degradation activity. the extracellular matrix, as well as by a decrease in the synthesis of the different glycosaminoglycans.

During aging of the skin, these changes usually result in the appearance of a greater microrelief of the skin or even fine lines, and finally the occurrence of deep wrinkles, a loss of elasticity, a touch rough, and a dry skin. Histologically, flattening of the dermoepidermal junction and a decrease in the thickness of the dermis and epidermis are observed.

In particular, skin hydration disorders, and in particular skin dryness, can often be observed with age.

Moreover, many external factors can further strengthen the drying of the skin or worsen the condition. These factors include climatic conditions such as cold or wind, sunlight, exposure to certain chemical or therapeutic agents.

Physiologically, dry skin is often associated with a decrease in the level of skin hydration and a change in the maturation process of the stratum corneum, with the most visible sign of dander on the skin surface. . On the sensory level, dry skin can be characterized by a feeling of tightness and / or cutaneous tension.

The collagen and glycosaminoglycan content of the skin is also decreased and the barrier function of the aged skin can be impaired.

Many epidermal factors, whose expression, biological activity or maturation are altered, diminished or increased, are known to be involved, directly or indirectly, in the occurrence and manifestation of the aged skin or the various signs of aging. associated skin, such as dry skin.

These factors can be used as bio-markers of the skin, as screening targets or even as cosmetic actives. However, there are still many unknowns as to the intimate mechanism and all the factors involved in the aging of the skin, associated or not with dehydration.

Thus, there remains a need to identify new biomarkers of the skin, and in particular able to characterize an aged skin or signs of skin aging, in particular aged and dry skin or signs of dry skin associated with an aged skin.

There is also a need for new biomarkers to characterize a state of the skin, and in particular an aged skin or signs of skin aging, particularly aged and dry skin or signs of dry skin associated with an aged skin. .

There is still a need for new tools for screening active agents or physical treatments that are suitable for skincare, and more particularly useful for preventing and / or treating aged skin or signs of skin aging, in particular a skin condition. old, dry skin or signs of skin dryness associated with aged skin.

There is still a need for new assets or treatments for the prevention and / or treatment of aged skin or signs of skin aging, especially aged and dry skin or signs of skin dryness associated with aged skin.

There is still a need for new cosmetic targets for the care of the skin, and in particular for the prevention and / or treatment of aged skin or signs of skin aging, in particular aged and dry skin or skin conditions. signs of skin dryness associated with aged skin.

There is also a need to identify new biomarkers of dry skin or signs of skin dryness.

There is also a need for new biomarkers to characterize dry skin or signs of skin dryness.

There is still a need for new tools or targets for the screening of active agents or physical treatments suitable for the care of dry skin, and more particularly useful for preventing and / or treating dry skin or signs of dry skin. . There is still a need for new tools or targets for the screening of active agents or physical treatments to promote or enhance the hydration of the skin.

There is still a need for new actives or treatments to promote and / or enhance the hydration of the skin.

There is still a need for new assets or treatments for the prevention and / or treatment of dry skin or signs of skin dryness.

There is still a need for new cosmetic targets for the care of the skin, in particular to promote and / or enhance the hydration of the skin and in particular for the prevention and / or treatment of dry skin or signs of skin. cutaneous dryness.

The object of the present invention is to satisfy these needs.

Thus, according to one of its first objects, the present invention relates to the use (i) of at least one amino acid sequence encoded by a nucleic acid sequence represented by SEQ ID NO: 1, an analogue or a fragment of said amino acid sequence, or (ii) said nucleic acid sequence, as a biomarker of a skin condition.

Preferably, a biomarker of the invention is a biomarker of aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin.

A biomarker of the invention may also be a biomarker of dry skin or cutaneous signs of dryness.

Thus, the use of (i) at least one amino acid sequence encoded by a nucleic acid sequence represented by SEQ ID NO: 1, an analogue or a fragment thereof, is also described. amino acid, or (ii) said nucleic acid sequence, as a biomarker of a state of hydration of the skin, and in particular of dry skin and / or signs of dryness of the skin.

For the purposes of the invention, the term "biomarker" means a molecule or the activity of a molecule, the presence, content or degree of activity of which is characteristic of a biological, physiological or pathological process, or impact or the effect induced by the administration of an active agent or a physical treatment on such a process.

For the purposes of the invention, the term "skin" refers to the entire epidemic of the human body, including the scalp and the lips. More particularly, the skin considered in the present invention is preferably the lips, the skin of the face, décolleté, arms or legs, and preferably the skin of the face and / or décolleté.

Unexpectedly, during a differential proteomics study using the "iTRAQ" technology, the inventors observed from proteins extracted from varnished strippings of populations of different age classes that Dermcidin (or DCD) was found to be a sensitive and specific biomarker of the aged skin.

More specifically, the inventors have observed that the level of expression of Dermcidin, and more particularly of peptides derived from Dermcidin and identified by the sequences SEQ ID NO: 17 and SEQ ID NO: 18, were systematically decreased in the aged skin by compared to young skin.

In addition, unexpectedly, in a Western blot comparative proteomics study, the inventors observed from varnished strippings extracted proteins from different age-class populations with different degrees of skin hydration than Dermcidin. (or DCD) was found to be a sensitive and specific biomarker of dry skin.

Thus, the inventors have observed that the level of expression of Dermcidine, and more particularly of a peptide identified by the sequence SEQ ID NO: 16, was systematically decreased in dry skin compared to the normally hydrated skin. To the inventors' knowledge, this cutaneous surface antimicrobial protein has never been classified as varying during aging or during dehydration of the skin and even less as one of the most relevant biomarkers of this type. studies.

It is known that sweating decreases with age (Anderson and Kenney 1987, Kenney and Fowler 1988) and that Dermcidin is a sweat protein (Schittek, Hipfel et al, 2001). On the other hand, the experimental data obtained by the inventors unexpectedly show that there is a specific deficit of expression and / or maturation of the DCD during cutaneous aging, because this is under-represented. in cutaneous samples, even compared to other proteins of sweat origin. In addition, it can be pointed out that dry skin is a complex phenomenon that is not systematically associated with a sweat deficit.

According to yet another of its objects, the present invention relates to the use (i) of at least one amino acid sequence of the invention, or (ii) a nucleic acid sequence of the invention, for screening for active agents or physical treatments capable of modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence.

The active agents or the screened physical treatments may be more particularly intended to prevent and / or treat the aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin.

Also described is a use (i) of at least one amino acid sequence of the invention, or (ii) a nucleic acid sequence of the invention, for screening for active agents or physical treatments which are capable of modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence and intended to promote and / or enhance the hydration of the skin.

Preferably, such active agents or physical treatments may be intended to prevent and / or treat dry skin and / or signs of dry skin.

For the purposes of the invention, the term "expression" with respect to an amino acid sequence, for example a protein or a peptide, or a nucleic acid sequence, for example an mRNA, its content or the variation of its content with respect to a reference.

For the purposes of the invention, the term "maturation" with respect to an amino acid sequence, for example a protein or a peptide, or a nucleic acid sequence, for example an mRNA, the modifications who follow their synthesis in a cellular environment. For example, in the case of an amino acid sequence, "maturation" is understood to mean post-translational modifications, such as the glycosylation or the farnesylation of certain amino acids, or the proteolytic steps leading to the elimination of sequences. so-called "signal" or "secretory" or the release of sequences with particular biological properties. In the case of a nucleic acid sequence, the term "maturation" is understood to mean, for example, the alternative splicing of an mRNA. Within the meaning of the invention, the term "activity", with respect to an amino acid sequence, for example a protein or a peptide, the biological activity of the amino acid sequence, where appropriate after maturation, such as enzymatic activity, receptor agonist or antagonist activity, or enzyme activator or inhibitor, so-called "structure" activity, or antimicrobial activity.

Within the meaning of the invention, the term "activity", with respect to a nucleic acid sequence, for example an AR m, its translation.

According to another of its objects, the present invention relates to the use (i) of at least one amino acid sequence of the invention, or (ii) at least one nucleic acid sequence of the invention , to characterize the effectiveness of a cosmetic treatment of the skin.

Still described is a use (i) of at least one amino acid sequence of the invention, or (ii) at least one nucleic acid sequence of the invention, to characterize the efficacy of a cosmetic treatment of dry skin and / or signs of skin dryness.

According to a preferred embodiment, a cosmetic treatment whose effectiveness is characterized may be a treatment of the aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin.

According to yet another of its objects, the present invention relates to the cosmetic use of an effective amount (i) of at least one amino acid sequence of the invention, or (ii) at least one sequence of nucleic acids of the invention, or (iii) at least one agent modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence, active agent for preventing and / or treating aged skin and / or cutaneous signs of aging, associated or not with dry skin.

Also described is a cosmetic use of an effective amount (i) of at least one amino acid sequence of the invention, or (ii) at least one nucleic acid sequence of the invention, or iii) at least one agent modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence, as active agent for promoting and / or reinforcing hydration of the skin. Preferably, such use may be dedicated to preventing and / or treating dry skin and / or signs of dry skin.

Within the meaning of the present invention, the term "effective amount" of a compound of the invention, a sufficient and necessary amount of this compound to obtain a desired effect, and more particularly a cosmetic effect or care with regard to old skin and / or cutaneous signs of aging, associated or not with dry skin. Also, an "effective amount" of a compound of the invention may be a sufficient and necessary amount of this compound to achieve a cosmetic or caring effect with respect to the hydration of the skin, and in particular the dry skin and / or signs of skin dryness.

For the purposes of the invention, the term "prevent" means reducing the risk of occurrence or slowing the occurrence of a given phenomenon, namely in the present invention, the aged skin and / or the signs of aging. cutaneous, associated (s) or not with a cutaneous dryness. Also, within the meaning of the invention, the term "prevent", reducing the risk of occurrence or slow onset of dry skin and / or signs of dry skin.

According to yet another of its objects, the present invention relates to the use of an effective amount (i) of at least one amino acid sequence of the invention, or (ii) at least one sequence of nucleic acids of the invention, or (iii) at least one modulating agent of the invention, for preparing a pluristratified epithelial cell model.

Preferably, a pluristratified epithelial cell model prepared according to the invention may be a reconstructed skin model.

According to yet another of its objects, the present invention relates to a method for characterizing a state of the aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin, comprising at least the steps consists in :

a) performing, in an isolated sample of a skin, a qualitative or quantitative measure of the expression, the maturation or the activity of said amino acid sequence or said nucleic acid sequence, and

b) comparing said measurement made in step a) with a reference measurement.

Depending on the difference observed between the measurement obtained and the reference measurement, the skin will be described as normally young skin or aged, even aged and dry skin. According to a preferred embodiment, a state of the skin considered in the invention is an aged skin selected from skins having undergone chronological aging and / or photo-induced aging.

Also described is a method for characterizing a state of hydration of the skin, and in particular dry skin and / or cutaneous signs of dryness, comprising at least the steps of:

b) comparing said measurement made in step a) with a reference measurement.

Depending on the difference observed between the measurement obtained and the reference measurement, the skin will be described as normally hydrated skin or dry skin.

According to yet another of its objects, the present invention relates to a method for screening active agents or physical treatments capable of modulating the activity, expression or maturation of an amino acid sequence of the invention or a nucleic acid sequence of the invention, comprising at least the steps of:

a) disposing said amino acid sequence or said nucleic acid sequence under conditions conducive to the activity, expression or maturation of said sequences,

b) contacting said amino acid sequence or said nucleic acid sequence with at least one active agent to be tested, or exposing said amino acid sequence or said nucleic acid sequence with a physical treatment to be tested,

c) performing a qualitative or quantitative measurement of the expression, the maturation or the activity of said amino acid sequence or said nucleic acid sequence, and

(d) compare that measure with a reference measure.

Depending on the difference observed between the measurement obtained and the reference measurement, the compound or the physical treatment screened will be characterized or not as being useful for preventing and / or treating the aged skin and / or the cutaneous signs of aging, associated with ) (s) or not to a dry skin. According to a preferred embodiment of the invention, the active agent (s) or the physical treatment (s) screened may be more particularly dedicated to preventing and / or treating aging skin and / or cutaneous signs of aging, associated (e) (s) or not to a dry skin.

Also described is a method of screening for active agents or physical treatments capable of modulating the activity, expression or processing of an amino acid sequence of the invention or a nucleic acid sequence of the invention and intended to promote and / or enhance the hydration of dry skin, comprising at least the steps of:

(d) compare that measure with a reference measure.

According to a preferred embodiment, a method or a use in accordance with the invention may be implemented in vivo, in vitro, or ex vivo, and even more preferably in vitro or ex vivo.

According to yet another embodiment, the present invention relates to an isolated peptide represented by an amino acid sequence selected from SEQ ID NO: 17 or SEQ ID NO: 18, an analogue or a fragment thereof.

The present invention has the advantage of proposing a new sensitive and specific biomarker of the skin, and in particular of the aged skin or signs of skin aging, associated (s) or not with a dry skin.

Also, the present invention has the advantage of providing a new biomarker sensitive and specific to the hydration of the skin, and in particular dry skin or signs of dry skin. The observation of the presence of Dermcidine in the stratum corneum, and more particularly of peptides derived specifically from this protein, advantageously makes it possible to determine quantitatively or qualitatively the expression or the activity of this protein, or the corresponding peptides. by simple topical sampling.

The sampling method may for example be a stripping type of technique to apply on the epidemic considered a portion of tape. By peeling off this tape, a fraction of the surface of the skin is removed. After protein extraction, this can then be analyzed by conventional methods, such as the enzyme-linked immunosorbent assay ELISA or Western-Blot analysis, or more particularly by the differential proteomic method iTRAQ as defined below.

Also, the present invention has the advantage of being able to provide a novel biomarker suitable for screening new active agents or new physical treatments suitable for the prevention and / or treatment of aged skin and / or cutaneous signs of aging, Associated (s) or not with a cutaneous dryness. Also, the present invention has the advantage of being able to provide a new biomarker suitable for screening new active agents or new physical treatments useful for promoting and / or enhancing the hydration of the skin. Preferably, such agents or treatment may be suitable for preventing and / or treating dry skin and / or signs of dry skin.

According to another advantage, the present invention makes it possible to propose new active agents that are suitable for the prevention and / or treatment of aged skins and / or cutaneous signs of aging, associated or not with a drought. skin.

Sequences of amino acids and nucleic acids

Dermcidine or Preproteolysin (DCD) is a protein of 110 amino acids (SEQ ID NO: 11) comprising a signal peptide of 19 amino acids (SEQ ID NO: 14) whose gene is located on chromosome 12, locus 12ql3.1 . Two isoforms of this protein are identified, one shorter, isoform 1 (SEQ ID NO: 12), the other longer, iso form 2 (SEQ ID NO: 13). It is secreted in sweat (Schittek, Hipfel et al., 2001). After cleavage of the signal peptide, Dermcidine (DCD) is a protein of 90 amino acids according to the sequence SEQ ID NO: 16.

After proteolytic post-translational processing, essentially by Cathepsin D, which takes place in sweat (Baechle, Flad et al., 2006), this protein precursor gives rise to DCD-1 (SEQ ID NO: 19) and DCD-IL (SEQ ID NO: 20) as well as many peptides ranging in length from 25 to 48 aa (Flad, Bogumil et al, 2002).

Unless otherwise indicated, the term "Dermcidin" is intended to mean in this application the amino acid sequences represented by SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 16, SEQ ID NO: 19 and SEQ ID NO: 20, whether or not undergoing post-translational processing.

According to a preferred embodiment, Dermcidine is more particularly referred to as the amino acid sequence represented by SEQ ID NO: 16.

Among the peptides derived from the maturation of Dermcidin, as described in US 2008/020976, some show antimicrobial activities against various microorganisms, such as Staphylococcus aureus, Escherichia coli, Enterococcus faecalis, or Candida albicans. The antimicrobial peptides are derived from the C-terminal portion of the protein.

The N-terminal part of the protein (SEQ ID NO: 15) consists of a bio-logically active factor promoting cell survival (Cunningham, Hodge et al., 1998).

This protein is constitutively expressed in the sweat gland (Rieg, Garb et al., 2004), and is implicated in particularly important PI3K / AKT / mTOR signaling pathways in cell proliferation and survival (Moreira, Strauss et al., 2008). The decrease in DCD observed in the sweat of individuals with atopy could partly explain their microbial susceptibility (Rieg, Steffen et al., 2005).

Some of the antimicrobial and N-terminal fragments are capable of stimulating the production of cytokines / chemokines by keratinocytes or other cell types involved in the regulation of cutaneous immunity (Watchorn, Dowidar et al., 2005). Niyonsaba, Suzuki et al., 2009). The use of DCD has recently been proposed as a particularly sensitive specific marker of presence of traces of sweat by RT-PCR or by ELISA usable in the forensic field (Sakurada, Akutsu et al., 2009).

According to one embodiment, an amino acid sequence suitable for the invention may be encoded by a nucleic acid sequence represented by SEQ ID NO: 1, or be an analogue or a fragment of this amino acid sequence.

By "analogue of an amino acid sequence" according to the invention is meant any amino acid sequence having a sequence identity of at least 85%, preferably at least 90%, and more. preferably at least 95% with said sequence, and a biological activity of the same nature.

By "biological activity of the same nature" with respect to an amino acid sequence according to the invention, one can hear the antimicrobial or stimulation properties of the survival and / or cell proliferation usually attributed to Dermcidin. Preferably, the term "biological activity of the same nature" with respect to an amino acid sequence according to the invention means the properties of prevention and / or treatment with regard to the aged skin or the signs skin aging, associated or not with a dry skin, or even with respect to dry skin and / or signs of skin dryness.

Sequence identity can be determined by visual comparison or by any computer tool commonly used in the domain, such as BLAST programs available at www.ncbi.nlm.nih.gov and used with default settings.

An analogue according to the invention may be a peptidomimetic agent. An analogue of an amino acid sequence of the invention may result from modifications resulting from mutation or variation in the sequences of the peptides according to the invention either from the deletion or from the insertion of one or more amino acids either the substitution of one or more amino acids or alternatively splicing. Many of these changes can be combined.

Advantageously, an analogue of an amino acid sequence of the invention may comprise conservative substitutions with respect to this amino acid sequence.

By way of example of mutations which may be considered in the present invention, it may be mentioned, in a non-exhaustive manner, the replacement of one or several amino acid residues by amino acid residues having a similar hydropathic index without substantially affecting the biological properties of the polypeptide. The hydropathic index is an index attributed to amino acids as a function of their hydrophobicity and charge (Kyte et al (1982), J. Mol Biol, 157: 105).

An amino acid sequence or an analogue thereof targeted by the present invention may be an amino acid sequence having undergone one or more post-translational processing (s).

By "post-translational processing (s)" is intended to encompass all the modifications that an amino acid sequence is likely to undergo at the end of its synthesis in a cell, such as, for example, a or phosphorylation (s), thiolation (s), acetylation (s), glycosylation (s), lipidation (s), such as farnesylation or palmitoylation, rearrangement Disulfide-type and / or cleavage-type structural bonding within the peptide sequence.

An analogue of an amino acid sequence has, moreover, substantially the same biological activity as this amino acid sequence.

It is moreover known that a primary amino acid sequence may comprise sites specifically recognized by protease-type enzymes, such as trypsin, which, once the recognition of these effective sites will induce the cleavage of the sequence by proteolysis. This proteolysis results in the generation of various peptides, or fragments of amino acid sequences of the invention.

Accordingly, the invention also extends to fragments of the

Dermcidin, possibly resulting from its proteolysis.

Within the meaning of the invention, the term "fragment of an amino acid sequence" means any portion of the amino acid sequence according to the invention comprising from 3 to 48 consecutive amino acids of said sequence, preferably from 6 to 36, preferably 8 to 32, and more preferably 10 to 30 consecutive amino acids of said sequence, and has a biological activity of the same kind.

According to one embodiment, an amino acid sequence that is suitable for the invention may be an amino acid sequence represented by a chosen sequence among SEQ ID NO: 11 to 20, especially among SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, an analogue, or a fragment thereof.

According to a preferred embodiment, an amino acid sequence that is suitable for the invention may be an amino acid sequence represented by a sequence chosen from SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, an analogue, or a fragment thereof.

Still more preferably, an amino acid sequence that is suitable for the invention may be an amino acid sequence represented by a sequence chosen from SEQ ID NO: 16, SEQ ID NO: 17, or SEQ ID NO: 18, SEQ. ID NO: 19 or SEQ ID NO: 20, an analogue or fragment thereof.

Preferably, an amino acid sequence that is suitable for the invention may be an amino acid sequence represented by a sequence chosen from SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19 or SEQ ID NO: 20, an analogue or a fragment thereof.

Even more preferably, an amino acid sequence suitable for the invention may be an amino acid sequence represented by a sequence chosen from SEQ ID NO: 17, SEQ ID NO: 18, an analogue or a fragment of that -this.

According to another embodiment, a polypeptide that is suitable for the invention may also be a natural or synthetic amino acid sequence, which may optionally be obtained after enzymatic or chemical lysis of Dermcidine or by chemical or biological synthesis, or by extraction from a biological tissue, such as the skin, naturally expressing this amino acid sequence or after transfection thereof, as well as the various post-translational forms thereof, or any natural or synthetic amino acids whose sequence completely or partially comprises a aforementioned amino acid sequence, for example variants and analogs.

Those skilled in the art can obtain an amino acid sequence according to the invention by means of recombinant DNA methods, for example those described in the manual "Molecular Cloning - A Laboratory Manual" (2nd edition). edition), Sambrook et al, 1989, Vol. I-III, Coldspring Harbor Laboratory, Coldspring Harbor Press, NY, (Sambrook). According to one object, the present invention relates as such to an isolated peptide represented by an amino acid sequence selected from SEQ ID NO: 17 or SEQ ID NO: 18, an analogue or a fragment thereof.

According to another embodiment, an amino acid sequence that is suitable for the invention may also be an amino acid sequence as defined above, fused with another amino acid sequence, a hydrophilic or hydrophobic targeting agent, a bio-conversion precursor, a luminescent, radioactive or colorimetric labeling agent.

Without limitation, examples of compounds that may be coupled to an amino acid sequence according to the invention include fluorescent proteins such as "Green Fluorescent Protein", fluorescent chemical compounds, such as rhodamine, fluorescein, or Texas Red ^® , compounds

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phosphors, radioactive elements, such as H, C, S, I or I, or colorimetric labeling agents such as chromogenic substrates sensitive to the action of galactosidase, peroxidase, chloramphenicol acetyltransferase, luciferase or alkaline phosphatase.

Depending on the nature of the compounds capable of being coupled with an amino acid sequence of the invention, the coupling may be carried out by chemical methods, in particular by means of reactive chemical functions or by molecular biological methods known to the art. skilled in the art.

Advantageously, an amino acid sequence of the invention may be encoded by a nucleic acid sequence selected from a sequence represented by SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, an analog or a fragment thereof.

According to one embodiment, the present invention also relates to nucleic acid sequences encoding an amino acid sequence of the invention and their implementation in the various uses and processes according to the invention.

Thus, the present invention also relates to a nucleic acid sequence, in particular of deoxyribonucleic acids, or of ribonucleic acids, represented by SEQ ID NO: 1, an analogue or a fragment thereof. For the purposes of the present invention, the term "nucleic acid sequence fragment" means a nucleic acid sequence comprising from 9 to 144 consecutive base pairs of said sequence, preferably from 18 to 108, preferably 24 to 96, and more preferably from 30 to 90 consecutive base pairs of said sequence, and encoding an amino acid sequence having a biological activity of the same nature as the amino acid sequence encoded by said sequence.

For the purposes of the present invention, the term "nucleic acid sequence analogue" means a nucleic acid sequence having a sequence identity of at least 85%, preferably at least 90%, and more. preferably at least 95% with said sequence, and coding for an amino acid sequence having a biological activity of the same nature as the amino acid sequence encoded by said sequence.

By "analog of a nucleic acid sequence" is meant a nucleic acid sequence, possibly resulting from the degeneracy of the nucleic acid code, and coding for an amino acid sequence according to the invention, in particular as defined above.

According to a preferred embodiment, a nucleic acid sequence of the invention may be represented by a sequence chosen from SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ. ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and SEQ ID NO: 10, an analogue or fragment thereof.

A nucleic acid sequence of the invention may be of any possible origin, namely either animal, in particular mammalian and even more particularly human, either plant, or micro-organisms, such as for example viruses, phages, or bacteria, among others, or fungi, without prejudging the fact that they are naturally present or not in said original organism.

According to one embodiment, the invention also relates to the isolated and purified nucleic acid sequences coding for an amino acid sequence considered according to the invention, as well as to their analogs and fragments.

A nucleic acid sequence according to the invention may comprise a sense, antisense or interferential sequence corresponding to a sequence coding for a polypeptide according to the invention. The subject of the invention is also nucleic acid sequences, in particular of ribonucleic or deoxyribonucleic acids, comprising a sense or antisense sequence, in particular "small interfering RNA" (siR A), corresponding at least to a sequence coding for a polypeptide of the invention or a nucleic acid sequence represented by SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6 , SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, an analogue or a fragment thereof.

Elderly skin and cutaneous signs of aging

By "aged skin" is meant a general condition of the skin resulting from chronological aging and / or photoinduced aging.

By "cutaneous signs of aging" is meant all changes in the external appearance of the skin due to aging, whether of chronological origin and / or photoinduced.

By way of example of these modifications considered in the invention, mention may be made of wrinkles and fine lines, wilted skin, lack of elasticity and / or tone of the skin, thinning of the dermis and / or degradation. collagen fibers, resulting in the appearance of soft, wrinkled skin.

It also includes all internal changes in the skin that do not always result in a modified external appearance, such as all internal skin damage, and more particularly the degradation of elastin fibers, or elastic fibers, consecutive to exposure to ultraviolet radiation.

In particular, the cutaneous signs of aging targeted by the invention are chosen from thinning of the skin, loss of firmness, loss of elasticity, loss of density, or loss of tone of the skin, dryness of the skin. skin, the appearance of a marked microrelief of the skin, the formation and / or the presence of fine lines and / or wrinkles, an alteration of the radiance of the complexion of the skin, a papery appearance of the skin, a Impaired skin odor, sagging skin, and wilting of the skin.

Preferably, the cutaneous signs of aging targeted by the invention are chosen from a thinning of the skin, the appearance of a microrelief of the skin marked, the formation and / or the presence of fine lines and / or wrinkles, sagging of the skin, and wilting of the skin.

In a still preferred manner, the cutaneous signs of aging targeted by the invention are chosen from the appearance of a microrelief of the marked skin, the formation and / or the presence of fine lines and / or wrinkles, a sagging of the skin. , and wilting of the skin.

The term "hydration of the skin" is understood to mean all the cellular and molecular mechanisms that lead to ensuring and maintaining the presence of a quantity of physiological water in the epidermis and the dermis, as well as the quantity of water resultant.

According to one aspect, the invention aims to maintain or even stimulate the homeostasis of these mechanisms, and thus to promote and / or enhance the hydration of the skin. Thus, in one aspect, the invention is directed to skins having a physiological hydrated state, also known as normal.

According to another aspect, the invention aims at restoring equilibrium or at reducing the risk of occurrence of an imbalance in the homeostasis of these mechanisms, and thus preventing and / or treating dry skin and / or signs of dryness. skin.

"Signs of skin dryness" means any changes in the appearance of the skin due to water deficiency in the epidermis and / or dermis.

Depending on the degree of manifestation of water deficit in the skin, dry skin may appear rough to the touch, wrinkled or even covered with scales. Dry skin can be manifested, for the most part, by a feeling of tightness and / or tension.

By "dry skin" is meant a general condition of the skin resulting from a deficit of water of the epidermis and / or dermis.

Dry skin may manifest as a desquamation disorder and present at different stages depending on the severity of this desquamation.

When the skin is slightly dry, these scales are abundant but not visible to the naked eye, the elimination is carried out by comeuocytes. They are less numerous but more and more visible to the naked eye when this disorder worsens, the clusters can include several hundred corneocytes, thus representing more or less large clusters, called dander.

Cutaneous dryness can be constitutional or acquired. In the case of constitutional dry skin, two categories can be distinguished: pathological skin and non-pathological skin.

The constitutional dry pathological skins are essentially represented by atopic dermatitis and ichthyoses. They are almost independent of external conditions and originate from known or unknown genetic modifications. Known genetic modifications affecting cutaneous hydration include, for example, modifications of the Transglutaminase-1 gene or those of the Filaggrin gene.

According to one embodiment, there is also described a pharmaceutical or dermatological composition comprising, in a physiologically acceptable medium, an effective amount (i) of at least one amino acid sequence of the invention, or (ii) at least one a nucleic acid sequence of the invention, or (iii) at least one agent modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence as an active agent for preventing and / or treating pathological constitutional dry skin selected from atopic dermatitis and ichthyosis.

In the case of non-pathological constitutional dry skin, the severity of the drought condition may, in turn, depend on external factors. Included in this skin category are senile skin (characterized by a general decrease in cutaneous metabolism with age), fragile skin (very sensitive to external factors and often accompanied by erythema and rosacea) and vulvar xerosis (d probable genetic origin and manifesting itself primarily on the face, limbs and back of the hands).

In the case of acquired dry skin, the intervention of external parameters such as exposure to chemical agents, difficult climatic conditions, solar rays or even certain therapeutic treatments (retinoids, for example) is crucial. Under these outside influences, the epidemic can become momentarily and locally dry. This can concern any type of epidemic.

Regardless of the origin, a skin with dryness of the skin may present, generally, the following signs: roughness to the touch and scaly, as well as reduced flexibility and elasticity. Dry skin, also called "xerosis", can appear at any age, and not be linked to a pathological condition. We will speak in this case of "acquired" drought.

However, xerosis becomes more common and troublesome with age, especially in women. This is called senile xerosis. In addition, women usually experience a worsening of skin dryness during menopause, probably due to the hormonal imbalance characteristic of this phenomenon. The most affected areas are the lower legs, the dorsal forearm and the hands.

As mentioned earlier, acquired drought can be influenced by external factors. For example, the appearance of dry skin can be promoted by cold, dry, and wintry weather. We are talking about winter xerosis. Skin dryness can also be induced by exogenous stress, of chemical origin, for example of the anionic detergent type, or else of mechanical origin (friction, shaving).

According to one embodiment, a cosmetic use of the invention may advantageously be suitable for preventing and / or treating senile or fragile dry skin, or xerosis, in particular chosen from xerosis vulgaris, senile xerosis, and winter xerosis.

Although no studies have shown that dryness has an impact on the origin and formation of fine lines and wrinkles that are mainly due to aging, on the visual side dry skin makes them more noticeable. Dry skin can therefore be associated with aged skin with wrinkles or fine lines.

In addition, on the sensory level, skin dryness is characterized by a feeling of tightness and / or itching. For obvious reasons, these manifestations are not only a source of discomfort, even pain, but also an unsightly appearance.

According to one embodiment, a cosmetic use that may advantageously be suitable for preventing and / or treating the feelings of tightness and / or itching associated with dry skin is also described.

By way of example of signs of cutaneous dryness considered in the invention, mention may be made of skin that is withered, lack of elasticity, suppleness and / or tone of the skin. the rough touch, the presence of crevices, desquamation, the presence of scales, or the wrinkles and fine lines associated with dry skin.

The signs of cutaneous dryness considered are also all internal changes in the skin that do not always result in a modified external appearance, such as, for example, all internal skin damage, and more particularly the degradation of elastin fibers, or elastic fibers, following drying of the dermis. According to a preferred aspect of the invention, the signs of cutaneous dryness are chosen from withered skin, a lack of elasticity, suppleness and / or tone of the skin, a rough feel, a presence of crevice, desquamation, a presence of scales, wrinkles and fine lines associated with dry skin, and a feeling of tightness and / or itching.

According to a very preferred aspect of the invention, the aged skin or the cutaneous signs of aging may or may not be associated with a dry skin. biomarker

The present invention relates to the use of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, as a biomarker of a skin condition.

Also described is the use of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, as a biomarker of a hydration state of the invention. skin, and more particularly dry skin and / or signs of skin dryness.

Preferably, a use according to the invention makes it possible to characterize a state of the skin such as the aged skin and / or the cutaneous signs of aging, associated or not with a dry skin.

According to one embodiment, a decrease in the activity, expression or maturation of said biomarker may be indicative of aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin.

According to one embodiment, it is also described that a decrease in the activity, expression or maturation of said biomarker may be indicative of skin deficient in hydration, and more particularly of dry skin. and / or signs of skin dryness. The present invention relates, according to another of its aspects, the use of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, to characterize the effectiveness of a cosmetic treatment of the skin, and to preference of the aged skin and / or cutaneous signs of aging, associated (s) or not with a cutaneous dryness.

Also described is the use of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, to characterize the effectiveness of a cosmetic treatment of the skin dry and / or signs of skin dryness.

According to one embodiment, an increase in the activity, expression or maturation of said biomarker may be indicative of an effective cosmetic treatment to exert a beneficial effect on the skin and more preferably an effect with respect to aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin.

It is also described that an increase in the activity, the expression or the maturation of said biomarker may be indicative of an effective cosmetic treatment to exert a beneficial effect on the skin in hydration deficit, and more preferably a effect on dry skin and / or signs of skin dryness.

In a use according to the invention, a decrease or an increase in the activity, the expression or the maturation of said biomarker can be determined by comparison with a reference measurement obtained according to any method known to man of art.

A "reference measurement" for a given parameter, is a qualitative or quantitative measure of this parameter carried out under so-called "control" or "normal" conditions, for example determined in a reference sample, or determined in a sample in the absence of a treatment alleged to have an effect on the parameter.

For example, a reference measurement for an amino acid sequence or a nucleic acid sequence according to the invention may be a quantitative or qualitative value relating to the expression, the maturation or the activity of said sequences determined in a skin sample young and physio logically healthy, or determined in a skin sample, especially aged skin, before a cosmetic treatment. Also for example, a reference measurement for an amino acid sequence or a nucleic acid sequence according to the invention may be a quantitative or qualitative value relating to the expression, the maturation or the activity of said sequences determined in a sample of physiologically healthy and normally hydrated skin, or determined in a sample of dry skin, before a cosmetic treatment.

Preferably, a reference measurement is a statistical measurement, that is to say having been repeated on different samples so as to obtain an average.

The reference measurement can be performed in parallel or sequentially with the test measurement.

It can also be a "historical" measure, that is to say carried out prior to the test measurement, and stored, for example in a database, for later use.

A comparison of the test measurement with a reference measurement, and an observation of a deviation or absence of deviation between the two measurements, makes it possible to derive information as to the measured parameter, for example the decrease or the increase. expression, processing or activity of an amino acid sequence or a nucleic acid sequence according to the invention.

Such information may be subsequently implemented to determine the youthfulness or age of a skin. Also, such information can be subsequently implemented to determine the normally hydrated or dry nature of a skin

A method according to the invention can also be carried out on a skin sample, taken from an epidermal cellular model, or a reconstructed isolated skin in order to qualify the state.

According to one embodiment, the invention relates to a method, in particular in vitro or ex vivo, for characterizing a state of the skin.

There is also described a method, in particular in vitro or ex vivo, for characterizing a state of hydration of the skin.

A method of the invention advantageously makes it possible to characterize an aged condition of the skin and / or signs of cutaneous aging, and more particularly to characterize an aged condition of the skin of chronological origin and / or photoinduced. It is also described that a method of the invention advantageously allows to characterize a dry state of the skin and / or signs of dry skin.

According to another embodiment, the invention relates to a cosmetic process, or non-therapeutic, in particular in vitro or ex vivo, to characterize the effectiveness of a cosmetic treatment of the skin, and preferably of the aged skin and / or or cutaneous signs of aging, associated or not with dry skin, in an individual in need, comprising at least the steps of:

a) performing, prior to the implementation of the cosmetic treatment, in a first isolated skin sample taken from said individual, at least a first qualitative or quantitative measure of the expression, the maturation or the activity of at least an amino acid sequence of the invention or at least one nucleic acid sequence of the invention,

b) performing, after the implementation of the cosmetic treatment, in a second isolated skin sample taken from said individual, at least a second measurement, qualitative or quantitative, of the expression, the maturation or the activity of said sequence amino acid of the invention or said nucleic acid sequence of the invention, and

c) comparing the first and second measurements, in particular to derive information relating to an effect at least of the implementation of the cosmetic treatment.

It goes without saying that the measurements made in steps a) and b) must be comparable to each other, and therefore be relative to the same parameter.

Preferably, a method of the invention makes it possible to demonstrate an effect of a cosmetic treatment capable of causing a regression of the aged skin and / or cutaneous signs of aging, associated (s) or not with a drought skin.

It is also described a cosmetic process, or non-therapeutic, in particular in vitro or ex vivo, to characterize the effectiveness of a cosmetic treatment of dry skin and / or signs of cutaneous dryness, in an individual in need, comprising at least the steps of:

a) performing, prior to the implementation of the cosmetic treatment, in a first isolated skin sample taken from said individual, at least a first qualitative or quantitative measurement of the expression, the maturation or the activity of a sequence of amino acids of the invention or a nucleic acid sequence of the invention, b) performing, after the implementation of the cosmetic treatment, in a second isolated skin sample taken from said individual, at least a second measurement, qualitative or quantitative, of the expression, the maturation or the activity of said sequence amino acid of the invention or said nucleic acid sequence of the invention, and

It goes without saying that the measurements made in steps a) and b) must be comparable to one another, and therefore be relative to the same parameter.

Preferably, it is also described that a method of the invention makes it possible to demonstrate an effect of a cosmetic treatment that can make the dry skin and / or the signs of dryness of the skin regress.

The qualitative or quantitative measurement of the expression, the maturation or the activity of a nucleic acid sequence of the invention can be determined by any method known to those skilled in the art.

By way of example of methods that are suitable for the invention, mention may be made of the polymerase chain reaction (PCR), quantitative (Q-PCR) or not, in the presence or absence of reverse transcriptase (RT-PCR or Q-RT-PCR), Northern-blot, ribonuclease protection assay, microarray methods, transcriptomic chip methods, oligonucleotide microarray methods, hybridization methods if you.

By way of example of agents suitable for the detection of a nucleic acid sequence of the invention, and in particular of an mRNA sequence, mention may be made of labeled nucleic acid probes which can be hybridizing to a nucleic acid sequence of the invention.

Such a nucleic acid probe can be easily obtained by any method known to those skilled in the art.

Thus, the nucleic acid sequences according to the invention can be used to produce sense and / or antisense oligonucleotide primers, which hybridize under conditions of high stringency to at least one of the sequences SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, an analogue or fragment thereof.

The expression of a nucleic acid sequence may also be determined, indirectly, by determining the expression of the amino acid sequence encoded by said sequence, by means of any technique known in the art, such as Western blot, ELISA, BRADFORD or LOWRY method, or as indicated below.

The qualitative or quantitative measurement, expression, maturation, or activity of an amino acid sequence of the invention may be performed by any method known to those skilled in the art.

As methods for detecting the expression, the maturation, or the activity of an amino acid sequence, mention may be made of Western blot, blot-blot, dot-blot, Single or multiplexed Enzyme Linked Immuno-Sorbent Assay (ELISA) methods, proteomics or glycomic methods, staining polypeptides in a silver-based dye polyacrylamide gel, Coomassie blue or SYPRO, immunofluorescence, UV absorption, immunohistochemical methods in classical electron or confocal microscopy, FRET (fluorescence resonance energy transfer), TR-FRET methods (time resolved FRET / FRET in time resolution), methods of fluorescence lifetime imaging microscopy (FLIM), FSPIM methods (fluorescence spectral imaging microscopy / imaging spectroscopy). fluorescence recovery after fluorescence recovery after photobleaching), reporter gene methods, atomic force microscopy (AFM) methods, surface plasmon resonance methods, fluorescence recovery after photobleaching (FRAP), microcalorimetry methods, flow cytometry methods, biosensor methods, radioimmunoassay (RIA) methods, isoelectric focusing methods, and enzymatic tests, methods using peptide chips, chips sugar, antibody chips, mass spectrometry methods, SELDI-TOF spectrometry methods (Ciphergen).

More generally, enzyme immunoassay methods from more quantitative and sensitive protein solutions can in particular be used. These ELISA methods combine couples of capture antibodies and specific detection of the targeted antigen. Specifically developed commercial or polyclonal, monoclonal or recombinant antibodies can be used. High capacity multiplexed ELISA techniques can also be implemented. It is thus possible to mention the Luminex antibody-type multiplexed approach (for example Bioplex from Bio-Rad), of the antibody-on-flat surface type ("antibodies-arrays") (for example, an approach proposed by MesoScale Discovery).

In particular, it may be advantageous to detect the expression of an amino acid sequence of the invention by means of an antibody, where appropriate in a labeled form. Such an antibody can be labeled with a directly detectable or detectable substance by reaction with another reagent.

By "antibodies" is generally meant monoclonal or polyclonal antibodies, as well as immunoglobulin fragments capable of binding an antigen and which may be produced by any genetic engineering technique known to those skilled in the art. by enzymatic or chemical cleavage of intact antibody.

An antibody that can be used as a tool for evaluating a state of an epidemic can be obtained by any method known to those skilled in the art, as described in "Antibodies: A Laboratory Manual" , Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1990).

According to a preferred embodiment, it may be advantageous to detect the expression of an amino acid sequence of the invention by means of a differential proteomic method "iTRAQ".

According to a preferred embodiment, it may also be advantageous to detect the expression of an amino acid sequence of the invention by means of a Western blot.

Such a method is known to those skilled in the art and can advantageously be implemented as described in the examples below.

In particular, the term "activity" with respect to an amino acid sequence of the invention means an antimicrobial activity, an activity for stimulating cell survival, an activity for stimulating cell proliferation, or a an activity for reducing or preventing aged skin and / or cutaneous signs of aging, associated or not with dry skin, as indicated above.

Also particularly, by "activity" with respect to an amino acid sequence of the invention is meant an antimicrobial activity, an activity for stimulating cell survival, an activity for stimulating cell proliferation, or an activity for reducing or preventing dry skin and / or signs of dry skin, as indicated above.

Such activity can be determined by any method known to those skilled in the art, such as, for example, by evaluating the proliferation or survival of epidermal cells in cultures, such as keratinocytes, or by evaluating the activity. antimicrobial on cultured bacteria, such as Staphylococcus aureus or Escherichia coll.

Preferably, the determination of a state of the skin or the characterization of the effectiveness of a cosmetic treatment of the skin can be carried out by measuring the variation of the expression of an amino acid sequence of the skin. invention, and preferably represented by a sequence selected from SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19 or SEQ ID NO: 20, an analogue or a fragment thereof. this.

The methods of the invention are particularly advantageous insofar as their implementation does not require the use of an invasive technique. An epidermis sample can thus be obtained by so-called "stripping" techniques and directly analyzed by a conventional analysis technique known to those skilled in the art.

These strippings are tacky surfaces applied to the surface of the epidemic such as Blenderm ^® 3M, D 'squam (commercial adhesive CuDERM), cyanoacrylate glue or the method of "stripping" varnish. Thanks to these "strippings", adherent corneocytes and the contents of their intercellular spaces can be removed and subsequently subjected to an extraction allowing access to the protein content.

Sampling of a sample suitable for a process of the invention may also be carried out more directly by "washing" the skin surface, for example by means of turbine-type propellers such as spiral cells. as described in the patent FR 2 667 778 associated with a fluid circuit, or simply by adding / removing a drop of buffer on the surface of the skin.

As an indication, other sampling methods adapted to the implementation of the invention may be mentioned, such as scraping methods of the upper part of the stratum corneum by means of a bimetallic system. This technique allows the collection of scales that can then be directly analyzed by different techniques to determine the levels of minerals, amino acids or lipids.

Advantageously, one of the markers of the invention can be used for the purpose of preclinical, more effective and more rigorous selection of individuals, with a view to evaluating the efficacy of treatment or of a cosmetic active ingredient for the care of the patient. the skin.

Also advantageously, one of the markers of the invention can be used for purposes of preclinical, more effective and more rigorous selection of individuals, in order to evaluate the effectiveness of treatment or cosmetic active for the care dry skin.

Also, a biomarker of the invention may advantageously be used as indicated above to evaluate the efficacy of an active agent, in vitro, ex vivo or in vivo with respect to a hydration deficit of the skin. , or dry skin or signs of dry skin.

Also, a biomarker of the invention may advantageously be used as indicated above to evaluate the efficacy of an active agent, in vitro, ex vivo or in vivo.

Also, a biomarker of the invention can be implemented to establish a personalized advice of a cosmetic treatment for an individual according to its expression profile of cutaneous biomarkers. Screening

According to one of its aspects, the present invention relates to the use of an amino acid sequence, of nucleic acids of the invention, for screening or in a screening method, in particular in vitro or ex vivo, of active agents or physical treatments, particularly suitable for the care of the skin. The active agents or the screened physical treatments may in particular be suitable for the care of the aged skin, and / or for the prevention and / or treatment of the cutaneous signs of aging, Associated (s) or not with a cutaneous dryness. The use of an amino acid sequence, of nucleic acids of the invention is also described for screening or in a screening method, in particular in vitro or ex vivo, of active agents or physical treatments. , particularly suitable for the care of the skin and intended to promote and / or enhance the hydration of the skin.

Also, the active agents or the screened physical treatments may in particular be suitable for the care of dry skin, and / or for the prevention and / or treatment of the signs of dry skin.

A use or a method of the invention may comprise comparing a measurement of the activity, expression or processing of an amino acid sequence or nucleic acid sequence according to the invention. invention to a reference measurement.

A reference measurement may be as previously defined.

In particular, a reference measurement may be a quantitative or qualitative value relative to the expression, the maturation or the activity of said sequences determined in a sample in the absence of active agent or physical treatment tested.

Thus, a reference measurement can be obtained by repeating the steps of a method of the invention, and in particular steps a), b) and c) of a method of the invention as defined above, in the absence of biological or chemical compounds, or physical treatments to be tested.

A comparison of the test measurement with a reference measurement, and an observation of a deviation or absence of deviation between the two measurements, makes it possible to draw information as to the effect of the active agent or the physical treatment. tested.

The quantitative or qualitative determination of the expression, the maturation or the activity of an amino acid sequence or of a nucleic acid sequence of the invention may be carried out by any method known to man of art, and especially as described above.

According to one embodiment, the screening of an active agent or a physical treatment capable of modulating the activity of an amino acid sequence of the invention can be done by measuring the activity or the expressing a target molecule belonging to the signaling or metabolism pathways in which may be involved said of an amino acid sequence, such as for example a reporter gene system. According to one embodiment, a method of the invention may be implemented in a cell-free system, ie in a system not comprising cells but reproducing cellular functions, or in an isolated cell sample.

A method according to the invention can be carried out on an isolated cell sample, an acellular sample, an isolated amino acid sequence or an isolated nucleic acid sequence of the invention, obtained by cutaneous biopsy, from cells in culture, in particular from an epidermal model, or a non-invasive skin surface sample, in particular by stratum corneum tape-stripping or by simple washing, as previously described.

Advantageously, as a cellular sample that is suitable for the invention, there may be mentioned a sample of keratinocytes or any other cell type of the skin expressing an amino acid sequence of the invention.

Preferably, the screening of an active agent or a physical treatment can be carried out by measuring the variation of the expression, in the presence and absence of the active agent or the screened physical treatment, of a sequence of amino acids of the invention, and preferably represented by a sequence selected from SEQ ID NO: 16, SEQ ID NO: 17, or SEQ ID NO: 18, an analogue or a fragment thereof.

Modulating agent

For the purposes of the present invention, the term "modulating agent" or

"Active agent or a physical treatment capable of modulating the expression, the maturation or the activity of an amino acid sequence or a nucleic acid sequence according to the invention", any compound or physical phenomenon capable of to act, directly or indirectly, on at least one amino acid sequence or a nucleic acid sequence according to the invention, or on an element of an intra- or extracellular signaling pathway, or a metabolic pathway, or regulation of transcription and / or translation involving said amino acid sequence or said nucleic acid sequence.

For the purposes of the invention, the term "modulate", with respect to a given effect, the action of stimulating or inhibiting this effect.

The active agents or the physical treatments resulting from a screening according to the invention can be advantageously used for cosmetic purposes, especially in aging skin or cutaneous signs of aging, associated or not with dry skin. It is also described that the active agents or the physical treatments resulting from a screening according to the invention can be advantageously used for cosmetic purposes, especially with regard to the hydration of the skin, and in particular of dry skin or skin. signs of skin dryness.

According to one embodiment, an active agent of the invention may be a stimulating agent for the activity, expression or maturation of an amino acid sequence of the invention.

In particular, a stimulating agent may be chosen from among a lysate of Bifidobacterium sp. and a mixture of a lysate of Bifidobacterium sp. and phytosphingosine-salicylate.

Preferably, a lysate of Bifidobacterium sp. may be a lysate of Bifidobacterium longum. A lysate of Bifidobacterium longum suitable for the invention may be the lysate registered under the name INCI: Bifidate ferment Lysate, under the name EINECS: Bifidobacterium longum, under the EINECS No.: No. 306-168-4, and under the N ° CAS: N ° 96507-89-0. Such a lysate is in particular sold under the name Repair Complex CLR ^® by the company K. RICHTER GmbH.

A mixture composed of a lysate of Bifidobacterium sp. and phytosphingosine-salicylate according to the invention may comprise a lysate of Bifidobacterium longum as defined above and a phytosphingosine-salicylate derivative.

A phytosphingosine salicylate derivative suitable for the invention may be derived sold by Goldschmidt under the name EVONICK phytosphingosine SLC ^®.

A mixture according to the invention may comprise a lysate of Bifidobacterium sp. at a rate of 10% and a phytosphingosine-salicylate derivative at a rate of 0.002%.

As a modulating agent that can be screened according to a use or a process in accordance with the invention, mention may also be made of antibodies or interfering RNAs. compositions

The present invention also relates to compositions, in particular cosmetic compositions, comprising in a physiological or cosmetically environment acceptable an effective amount of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, or at least one active agent capable of modulating the activity, expressing or processing said amino acid sequence or said nucleic acid sequence.

More particularly, the invention relates to a cosmetic composition comprising a peptide represented by an amino acid sequence selected from SEQ ID NO: 17 or SEQ ID NO: 18, an analogue or a fragment thereof or an acid sequence. nucleic acid coding for such a peptide.

For the purposes of the present invention, the term "physiologically acceptable medium" is intended to mean a medium which is suitable for the administration of a composition topically to the skin, the scalp or the lips, orally or via parenteral route such as the intradermal or subcutaneous route.

A composition of the invention may contain adjuvants customary in the field in question, such as hydrophilic or lipophilic gelling agents, hydrophilic or lipophilic additives, preservatives, antioxidants, solvents, perfumes, fillers, filters, absorbers odor and coloring matter.

The amounts of the various constituents of the compositions according to the invention are those conventionally used in the fields under consideration.

The amount of amino acid sequence, or nucleic acid sequence of the invention, or active agent according to the invention contained in a composition of the invention, also called "effective amount" is, of course depending on the nature of the asset and the effect sought and may therefore vary to a large extent.

To give an order of magnitude, a composition may contain an amino acid sequence, or a nucleic acid sequence, or an active agent according to the invention in an amount representing from 0.00001% to 50% of the total weight of the composition, in particular in an amount representing from 0.001% to 10% of the total weight of the composition, and more particularly in an amount representing from 0.1% to 1% of the total weight of the composition.

According to another embodiment, a cosmetic composition of the invention may comprise, in addition, at least one additional active cosmetic and / or therapeutic agent. Additional assets

As examples of additional active agents that may be used in the context of the present invention, mention may be made of cosmetic oils, such as silicone oils, vegetable oils of the triglyceride type, and hydrocarbon oils such as PARLEAM oil. and esters of fatty acids and fatty alcohol.

It may also be possible to use other active agents that make it possible to improve the state of the skin and / or its annexes, such as moisturizing or moisturizing active agents or active agents that make it possible to improve the natural lipid barrier, such as ceramides, cholesterol sulphates and / or fatty acids, and mixtures thereof.

It may also be possible to use other active agents which make it possible to improve the state of the skin and / or its appendages, such as anti-aging active agents.

It may also be possible to use enzymes having an activity on the skin and / or its annexes, such as proteases, lipases, glucosidases, amidases, cerebrosidases and / or melanases, and mixtures thereof.

Other examples of active agents that are suitable for the implementation of the present invention include: analgesic active agents, anti-yeast active agents, antibacterial active agents, antiparasitic active agents, antifungal active agents, antiviral active agents, anti-inflammatory active agents steroidal agents, anesthetic active agents, antipruritic active ingredients, keratolytic active agents, anti-free radical active agents, anti-seborrhoeic active ingredients, anti-dandruff active ingredients, anti-acne active agents, active agents to prevent skin aging and / or to improve its condition, anti-dermatitis active agents, anti-irritant active ingredients, immunomodulatory active ingredients, active agents for the treatment of dry skin, antiperspirant active ingredients, anti-psoriatic active ingredients, protective active ingredients against UV, antihistaminic active ingredients, healing active ingredients, auto-broning active ingredients, antioxidants such as green tea or active fractions thereof, glycerine, laponite, caffeine, aromatic essential oils, dyes, depigmenting active ingredients, lipo-regulators, softening, refreshing, deodorant, numbing, whitening, nourishing active ingredients, agents that reduce differentiation and / or proliferation and / or skin pigmentation, and mixtures thereof.

As additional active agents that may be more particularly suitable for the invention, mention may also be made of probiotic microorganisms, other than those considered in the previously described lysate, prebiotic active agents, active ingredients promoting the synthesis of skin defense factor, active agents that restore the equilibrium differentiation / proliferation of the cells of the epidemic, in particular such as retinol-like or retinol-like active agents, moisturizing active agents or stimulating active ingredients cutaneous protease activity, in particular Cathépsine D.

Cosmetic use

The present invention relates to the cosmetic use of an effective amount of at least one amino acid or nucleic acid sequence of the invention or at least one agent modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence, and in particular a modulating agent as defined above, as active agent for preventing and / or treating aged skin and / or cutaneous signs aging, associated (s) or not with a skin dryness.

It is also described the cosmetic use of an effective amount of at least one amino acid or nucleic acid sequence of the invention or at least one agent modulating the activity, the expression or maturation of said amino acid sequence or said nucleic acid sequence, and in particular a modulating agent as defined above, as an active agent for promoting and / or enhancing the hydration of the skin.

It is further described that a use of the invention may be dedicated to preventing and / or treating dry skin and / or signs of skin dryness.

According to another aspect, the present invention relates to a cosmetic process, or non-therapeutic, for preventing and / or treating aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin in a an individual in need thereof, the method comprising at least one step of administering to said individual at least one composition comprising as active agent (i) at least one amino acid sequence of the invention or (ii) at least one a nucleic acid sequence of the invention, or (iii) at least one agent modulating the activity, expression or maturation of said sequences of the invention, in particular as defined above.

A method or a use of the invention makes it possible to prevent and / or treat cutaneous aging, in particular chronological and / or photoinduced aging. A method or a use of the invention makes it possible to confer and / or restore a youthful appearance to the skin, manifesting itself in a smooth, elastic, toned or firm skin, or a luminous complexion.

A method or a use of the invention makes it possible to prevent and / or reduce the presence of a marked microrelief of the skin, in particular at the commissure of the lips and crow's feet.

A method or use of the invention enhances the barrier properties of the skin.

It is also described a cosmetic or non-therapeutic method for promoting and / or strengthening the hydration of the skin, and preferably for preventing and / or treating dry skin and / or signs of dry skin, in an individual in wherein the process comprises at least one step of administering to said individual at least one composition comprising as active agent (i) at least one amino acid sequence of the invention or (ii) at least one sequence nucleic acids of the invention, or (iii) at least one agent modulating the activity, expression or maturation of said sequences of the invention, in particular as defined above.

A method or a use of the invention also makes it possible to confer and / or restore a hydrated and healthy appearance to the skin, manifesting itself in a smooth, elastic, toned or firm skin, or a luminous complexion.

A method or a use of the invention also makes it possible to prevent and / or reduce the presence of scales, roughnesses, cracks, flakes and / or wrinkles or fine lines associated with dry skin or lack of hydration. .

A method or use of the invention also enhances the barrier properties of the skin.

Preferably, a method of the invention may comprise the topical application to at least a portion of the skin of an individual in need thereof, in particular on the skin of the face and / or décolleté, from least one layer of a topical composition of the invention.

Advantageously, a method of the invention topically may comprise the application on the skin, and in particular on the skin of the face, of a composition of the invention in the form of a mask. A topical cosmetic process according to the invention may advantageously comprise the application of a composition of the invention, in combination simultaneously, sequentially or separately in time with an additional cosmetic or dermatological composition that is distinct from the composition of the composition. invention and intended for care and / or make-up of the skin.

According to another preferred embodiment, a cosmetic process of the invention may be carried out orally, in particular by administering at least one food or dietary composition for cosmetic purposes.

According to another preferred embodiment, a cosmetic process of the invention can be implemented parenterally. The parenteral implementation of a cosmetic process of the invention is carried out excluding any surgical procedure and is intended only to exert a surface treatment of the skin for aesthetic purposes.

Thus, a cosmetic method of the invention parenterally is implemented by any injection technique or device suitable for intra- epidermal injection and / or intradermal and / or subcutaneous.

Such administration may be effected, for example, by mesotherapy. A cosmetic process parenterally is therefore translated only by a superficial break in the skin and therefore goes out of any medical or therapeutic setting.

For the parenteral route, alternatively, systemic patch administration may be preferred.

A cosmetic process according to the invention can be implemented on a daily basis, for example on the basis of a single administration per day, or of a divided administration in two or three times a day, for example once in the morning and once in the morning. times in the evening.

A cosmetic process according to the invention can be implemented over a period of time ranging from one week to several weeks, or even several months, this period being able to be repeated after periods of non-treatment for several months, or even years. .

By way of example, a cosmetic process according to the invention may be provided for administering a composition of the invention, for example, at a rate of 1, 2 or 3 times per day, or more, and generally for an extended period of at least 4 weeks, or even 4 to 15 weeks, with one or more periods of interruption if necessary.

Reconstructed skin

In another aspect, the present invention relates to the use of an effective amount of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, or at least one modulating agent of the invention for preparing a pluristratified epithelial cell model, preferably a reconstructed skin model.

The interest of developing organotypic models closest to in-vivo conditions is great for the evaluation of safety and efficacy of active ingredients and formulas for cosmetic and dermatological applications.

The use of at least one amino acid sequence of the invention, or at least one nucleic acid sequence of the invention, or at least one modulating agent of the invention in a medium of culture is likely to improve the quality of the models developed.

A reconstructed skin model according to the invention may be useful for mimicking elderly skin, possibly associated with cutaneous dryness, or conditions for restoring the homeostasis of an aged skin, possibly associated with cutaneous dryness.

In another aspect, the present invention relates to a method for preparing an isolated pluristratified epithelial cell model, and preferably an isolated reconstructed skin comprising, at least the step of contacting at least one effective amount of at least one amino acid sequence, or at least one nucleic acid sequence, or at least one modulating agent according to the invention with cells capable of generating isolated reconstructed skin, and in particular keratinocytes.

A reconstructed skin model can include different cell types, such as keratinocytes, fibroblasts, Langerhans cells and melanocytes. The fibroblast type cells may be irradiated or not.

Such models and their preparation are known to those skilled in the art. According to yet another aspect, the present invention also relates to a method for preparing a pluristratified epithelial cell model, preferably a reconstructed skin model, comprising at least one step of culturing cells of at least one cell type of said cell. model, said cells having been genetically modified to suppress the expression of an amino acid sequence or a nucleic acid sequence of the invention.

Obtaining cells genetically modified to suppress the expression of an amino acid sequence or of a nucleic acid sequence of the invention, called "knockout" cells, can be carried out by any known method of the skilled person.

By way of example, such cells can be obtained by transfection and homologous recombination of a nucleic acid fragment that is inserted into or takes the place of the gene expressing the amino acid sequence whose expression is to be deleted. .

Also, it may be possible to suppress the expression of a given gene by transfection into the cell of a nucleic acid sequence coding for a mRNA-specific interfering RNA derived from the gene whose expression is to be deleted.

Legends of the figures

Figure 1: represents the variation of the expression of Dermcidin (average ± sem) measured by Western-Blot according to the degree of hydration of the skin.

For the purposes of the present invention, "a" must be understood, unless otherwise indicated, in the sense of "at least one".

The examples and figures below are presented for illustrative and not limiting of the invention. EXAMPLES

Example 1

Expression of Dermcidin in the skin

1- Materials and methods

a- iTRAQ protocol

The detection of the expression of Dermcidin (DCD) was carried out using the iTRAQ method (Isobaric Tagging Reagents for Quantitative Proteomic Analysis) developed by Applied Biosystems. This method allows the quantification of peptides previously labeled by an isobaric tag. There are 4 different tags and up to 4 different types of samples can be compared in the same LC / MS-MS analysis.

The principle of this method is as follows:

- Digestion of protein extracts and labeling:

First, each protein extract is digested with a proteolytic enzyme: trypsin.

In a second step, each peptide mixture obtained is labeled with an isobaric tag having a total mass 145, comprising a "reporter" group of mass 114, 115, 116 or 117, and a mass "balance" group 31, 30. , 29 or 28, linked together by an M fragmentation site.

The labeling is obtained by reacting the Peptide Reactive Group of each tag with the primary amines of the peptides (N-terminal end of the peptides or the side chain of lysines or tyrosines.

- LC / MS-MS analysis and quantification:

After the labeling step, the samples are pooled and analyzed by mass spectrometry. A given peptide, present in several samples, will have an identical parent mass but its MS / MS fragmentation spectrum will generate the mass of the reporter group characteristic of each of the iTRAQ reagents. The comparison of the intensity of each reporter ion (114, 115, 116 or 117) then allows the quantification of the peptide.

The iTRAQ method is more specifically described by Zieske (J. Bot., 2006, 57: 1501) or Wiese et al. (Proteomics, 2007, 7: 340). b- Collection and treatment of skin samples

Stripping-varnish samples of stratum corneum with a surface area of 90 cm ² are obtained according to the protocol described by Mehul et al., J. Biol. Chem., 2000, 275 (17): 12841 -7, at the outer side of the leg. Two groups of subjects are taken:

1) "Young Skin" group: 18-40 years, 27 male subjects;

2) "Aged Skin" group: over 60, 35 male subjects.

These skin samples are then analyzed by proteomics using the so-called "isobaric labeling" technique, previously described, using the Appliance Biosystems iTRAQ Reagents Multiplex kit (4352135), in accordance with the manufacturer's instructions.

2- Results

Among the identified proteins, the peptides represented by the sequences SEQ ID NO: 17 and SEQ ID NO: 18 derived from Dermcidine (Accession No. P81605 - Ref .: Swissprot / Uniprot) are detected and quantified in four experiments.

The quantification results of Dermcidin are given in the following table:

These results show that the average aged / young iTRAQ ratio is equal to 0.37 ± 0.09. This measure reflects a strong expression of Dermcidin in the skins of the group "Young skin", and a sharp decrease in its expression in the skins of the group "Aged skin".

Dermcidin therefore proves to be an interesting and specific biomarker of the skin, and more particularly of an elderly skin condition, associated or not with a dry skin. Expression of Apolipoprotein D and Cathepsin D heavy chain was measured as comparative data.

The results obtained are expressed in the following table.

In comparison with Dermcidin, Apolipoprote- me D and the heavy chain of Cathepsin D are only marginally under-expressed in the "aged skin" group compared to the "young skin" group.

This study validates the use of Dermcidine as an exceptional biomarker of the skin, and more particularly of the aged skin, associated or not with a cutaneous dryness.

Example 2

Expression of Dermcidin in the skin

1- Materials and methods

a- Western-Blot Protocol

The detection of Dermcidin expression (DCD) was performed using Western-Blot method.

The principle of this method is as follows: the proteins are separated by electrophoretic migration on a 10-20% SDS-PAGE criterion gel (Bio-Rad). After a semi-dry transfer on a PVDF membrane, the proteins are incubated in a blocking solution (TBS + 0.1% Tween + 1% milk) and then incubated with the primary antibody directed against the protein of interest overnight at 4 ° C. A second incubation with the secondary antibody (peroxidase coupled) directed against the primary antibody is performed for one hour at room temperature.

Depending on the degree of sensitivity, different kits are used for revelation: ECL, ECL + or ECL Advance (Amersham). The image is acquired with FluorSmax (Biorad) and quantified bands using Quantity-One software (Biorad)

b- Collection and treatment of skin samples

120 females, aged 18 to 70, were selected for this study.

Four zones per leg have been delineated (external / internal left leg, external / internal right leg). The samples are taken by Stripping Varnish (lower leg site) according to the protocol described by Mehul et al., (J. Biol Chem., 2000, 275 (17): 12841-7).

For this study, the analysis is performed on an area: external left leg. Samples from varnished strippings were used for Western Blot analysis. Before the samples were taken, the skin types were characterized by clinical scoring.

The clinical scoring is done by a dermatologist according to the "Atlas of dry skin on the legs" to identify the different types of skin according to the degrees below:

0: normal skin, regular skin relief, smooth appearance

1: slightly dry skin, ribbed skin texture, rough appearance

2: dry skin, ribbed skin and some scales, rough appearance 3: very dry skin, many scales and some scales, rough appearance

4: extremely dry skin, many scales, rough appearance

Preparation of acetone powders

Each polishing stripping of 75cm ² is immersed in 75ml of acetone. The varnish dissolves and the comeuocytes are in suspension. The mixture is filtered through vacuum pump on 40μιη nylon membrane to collect comeuocytes. Two successive washes are performed with the same volume of acetone. Acetonitrile powders of stratum corneum are obtained in dry form. Extraction of samples

Two types of extraction are carried out successively: native extraction then denaturing.

Native extraction

The acetone powders are weighed and put into 1.5 ml Eppendorf tubes (Trefflab cat.No.96.07246.9.01). 100 μl of PBS 0.1% triton X100 buffer are added to 1 mg of acetone powder and the mixture is left for one hour in contact with ice. Then it is ground for 30 seconds in the ice. The medium is then recovered in Millipore Ultraree columns of 0.45 μιη and then centrifuged for 5 min at 10,000 g at 4 ° C.: the supernatant is then collected. A protein assay is performed according to the BCA technique (BCA Pierce kit). The samples are adjusted to a concentration of 0.15 mg / ml in Laemmli 4X buffer and stored at -20 ° C pending analysis.

Denaturing Extraction It is made from the base of the native extract. In 1.5 ml Eppendorf tubes (Trefflab cat.No.96.07246.9.01) ΙΟΟμΙ of Laemmli IX buffer is added to the pellet for 1 mg of acetone powder. The mixture is boiled for 10 min at 90 ° C and then milled for 20 seconds and centrifuged for 10 min at 10000 g / min. The supernatant is collected. A protein assay is performed according to the Bradford technique. Samples are adjusted to 1 mg / ml and stored at -20 ° C pending analysis.

These skin samples are then analyzed by Western-Blot as previously described. The results were obtained from the denatured extracts.

Statistical Analyzes

The ef and cutaneous dryness (clinical score) on the level of expression of bio-markers was analyzed both graphically by representing the mean values with their confidence interval according to age classes or clinical score cutaneous dryness, and using a multiple regression model with age and cutaneous dryness score parameters in covariates.

The significance of the coefficients (Student's test) associated with covariates makes it possible to test the association between the bio-marker studied with age and the drought score. Taking into account the two parameters in the model makes it possible to estimate the effect of one of the parameters while adjusting on the effect of the second (everything else being equal).

The level of test significance is set at 5% in an unadjusted bilateral approach. The analyzes were carried out using SPPSS software version 17. Expression levels of bio-markers were previously transformed (square root or Log10) when it was useful to both symmetrize their distribution and stabilize their variance.

2- Results

The results obtained, illustrated in Figure 1, clearly show a decrease in the expression of Dermcidine correlated with an increase in the degree of dehydration, or drought, of the epidemic. Statistical analysis confirms a significant decrease in the expression of Dermcidine in dry skin compared to normal skin (p = 0.046).

This study validates the use of Dermcidine as an exceptional biomarker of skin hydration, especially dry skin.

Example 3

Effect of a lysate of Bifidobacterium sp. on the expression of Dermcidin

1- Materials and methods

Samples of skin samples, and measurements of the expression of the

Dermcidin are carried out as indicated in Example 1.

Volunteers are female subjects aged between 30 and 65 years old. They were combined and divided into two subgroups according to the treatment administered, a "Placebo" group, and a "Treaty" group.

The group "Treaty" receives topically, on the leg, at a rate of 2 times a day, morning and evening, for 56 days, or 2 months a composition comprising 10% Repair Complex ^® in a formula corresponding support to oil emulsion in demineralised water Arlacel / Myrj containing 5% Parleam, 15% cyclopentasiloxane, 3% glycerin and 2% petrolatum.

The Repair Complex ^® comprises a lysate of Bifidobacterium longum registered under the name INCI: Bifida ferment Lysate, under the name EINECS: Bifidobacterium longum, under the N ° EINECS: N ° 306-168-4, and under the N ° CAS: N 96507-89-0. This lysate is sold under the name Repair Complex CLR ^® by the company K. RICHTER GmbH.

On the other hand, the "Placebo" group receives only the support formula.

For each subject condition "treated" and "placebo", an average of the ratios J56 / J0 is obtained, thus making it possible to evaluate the effect of the treatment.

The ratios J56 / J0 are calculated according to the following formula:

% treatment effect = 100 * J56 / J0 "treated" ratio - J56 / J0 "placebo" ratio

J56 / J0 "Placebo" ratio The raw data obtained from the analysis are as follows

The average value of the J56 / J0 "placebo" ratio is 1.85, while the average value of the J56 / J0 "treated" ratio is 2.36.

Treatment with Repair Complex® results in a 27% increase in signal compared to placebo.

Thus the topical administration of a Bifidobacterium longum lysate increases the expression of Dermcidine by approximately 30% after 2 months of treatment.

Topical administration of a lysate of Bifidobacterium sp. Thus, it is possible to restore a level of Dermcidin expression in the skin, and in particular of the aged skin, associated or not with a dry skin, which is conducive to reinforcing its protective action on the skin, reinforcing the barrier properties of the skin. skin, and reduce the signs of skin aging, associated or not with a dry skin.

Topical administration of a lysate of Bifidobacterium sp. It also restores a level of expression of Dermcidin in dry skin, which helps to reinforce its skin protection action, to reinforce the skin's barrier properties, and to reduce the signs of dryness of the skin.

This study also validates the use of Dermcidin as a biomarker to evaluate the efficacy of a cosmetic treatment of aged skin, as well as a screening tool for new active agents for the treatment of skin. elderly, with or without dryness.

Example 4

Effect of a lysate mixture of Bifidobacterium longum and phytosphingosine on the expression of Dermcidine 1- Materials and methods

Samples of skin samples, and measurements of the expression of Dermcidine are carried out as indicated in Example 1.

Volunteers are female subjects aged 40 to 45 years old. Each subject is his own control, both arms are removed but only one is treated with the tested product and the second arm receives a placebo.

The arm "Treaty" receives topically at a rate of 2 times a day, morning and evening, for 56 days, or 2 months a composition comprising 10% Repair Complex ^® and 0.002% phytosphingosine-SLC in a support formula corresponding to an oil / water emulsion demineralized Arlacel / Myrj containing 5% Parleam, 15% cyclopentasiloxane, 3% glycerine and 2% petrolatum.

The Repair Complex ^® contains a Bifidobacterium longum registered under the name INCI: Bifida ferment Lysate, under the name EINECS: Bifidobacterium longum, under the EINECS N °: 306-168-4, and under the CAS N °: 96507 -89 to 0. This lysate is sold under the name Repair Complex CLR ^® by the company K. RICHTER GmbH.

The phytosphingosine salicylate derivative is that sold by Goldschmidt under the name EVONICK phytosphingosine SLC ^®.

The "Placebo" arm receives only the support formula.

For each "Treaty" and "Placebo" condition, an average of the ratios J56 / J0 is obtained as indicated above, thus making it possible to evaluate the effect of the treatment. 2- Results

The raw data obtained at the end of the experiment are as follows:

Experience # 1

Ratio J56 / J0 Ratio J56 / J0

Placebo Treaty

0.75 1.66

Experience n ° 2

Ratio J56 / J0 Ratio J56 / J0

Placebo Treaty

0.45 0.81 The average value of the J56 / J0 "Placebo" ratio is 0.6, while the average value of the J56 / J0 "Treaty" ratio is 1.265.

The treatment effect is equal to 105%, and reflects a doubling of the expression of Dermcidine compared to "placebo".

Thus the topical administration of a mixture comprising a Bifidobacterium longuni lysate and phytosphingosine salicylate increases the expression of Dermcidin by more than 100%.

The topical administration of a mixture comprising a lysate of Bifidobacterium longum and phytosphingosine-salicylate thus makes it possible to restore a level of expression of Dermcidin in the skin, and especially associated aged skin. or not to a cutaneous dryness, favorable to reinforce its action of protection of the skin, to reinforce the barrier properties of the skin, and to reduce the cutaneous signs of the aging, associated or not with a cutaneous dryness. The topical administration of a mixture comprising a Bifidobacterium longum lysate and phytosphingosine-salicylate also makes it possible to restore a level of expression of Dermcidin in the skin, and in particular to dry, favorable skin. to reinforce its skin protection action, to reinforce the skin's barrier properties, and to reduce the signs of skin dryness.

BIBLIOGRAPHY

Anderson, R.K. and W. L. Kenney (1987). "Effect of age on heat-activated sweat glans." Physiol. 63 (3): 1089-1094.

Baechle, D., Flad, T., et al. (2006). "Cathepsin D is present in human eccrine sweat and involved in the postsecretory processing of the antimicrobial peptide DCD-IL" J. Biol. Chem. 281 (9): 5406-5415.

Cunningham, T.J., L. Hodge et al. (1998) "Identification of a survival-promoting peptide in medium conditioned by oxidatively stressed cells of nervous system origin" J. Neurosci 18 (18): 7047-7060.

Flad, T., Bogumil R. et al. (2002) "Detection of dermcidin-derived peptides in sweat by ProteinChip Technology. J. Immunol. Methods 270 (1): 53-62.

Kenney, W. L. and S. R. Fowler (1988) "Methylcholine-activated eccrine sweat gland density and output as a function of age" J. Appl. Physiol. 65 (3): 1082-1086.

Kyte et al., (1982) J. Mol. Biol 157: 105.

Mehul B. et al, (2000) "Identification and Cloning of a New Calmodulin-like Protein from Human Epidermis" J. Biol. Chem. 275 (17): 12841-12847

Moreira, D. F., B. E. Strauss et al. (2008) "Genes up- and down-regulated by dermcidin in breast cancer: a microarray analysis" Genet. Mol. Res. 7 (3): 925-932.

Niyonsaba, F. A. Suzuki et al. (2009) "The human antimicrobial peptide dermcidin activates normal human keratinocytes" Br. J. Dermatol. 160 (2): 243-249.

Rieg, S.C., Garbe et al. (2004) "Dermcidin is constitutively produced by eccrine sweat gland and is not induced in epidermal cells under inflammatory skin conditions" Br. J. Dermatol. 151 (3): 534-539.

Rieg S., Steffen II et al. (2005) "Defeciency of dermcidin-derived antimicrobial peptides in patients with atopic dermatitis correlates with an impaired innate defense of human skin in vivo" J. Immunol. 174 (12): 8003-8010.

Sakurada K., T. Akutsu, et al, (2009) "Detection of dermidin for sweat identification by real-time RT-PCR and ELISA" Forensic. Sci. Int. 2010 Jan. 30, 194 (1-3): 80-4. Epub 2009 Nov. 13. Sambrook et al., 1989, Vol. I-III, Coldspring Harbor Laboratory, Coldspring Harbor Press, NY.

Schittek, B, R. Hipfel et al. (2001): "Dermcidin: a novel human antibiotic secret peptide by sweat tassels" Nat. Immunol. 2 (12): 1133-11137.

Watchorn, T. M. N. Dowidar et al. (2005) "The chachectic mediator proteolysis inducing factor activates NF-kappaB and STAT3 in human Kupffer cells and monocytes" Int. J. Oncol. 27 (4): 1105-1111.

Wiese et al., (2007) Protein labeling by iTRAQ: A new tool for quantitative mass spectrometry in proteome research Proteomics 7 (3): 340-350

Zieske et al. (2006) "A perspective on the use of iTRAQ reagent technology for protein complex and proflling studies" J. Exp. Bot. 57 (7): 1051-1058.

Claims

Use of (i) at least one amino acid sequence encoded by a nucleic acid sequence represented by SEQ ID NO: 1, an analogue or a fragment of said amino acid sequence, or (ii) said nucleic acid sequence, as a biomarker of a state of the aged skin and / or cutaneous signs of aging, associated or not with a dry skin.

2. Use according to the preceding claim, wherein said nucleic acid sequence is represented by a sequence selected from SEQ ID NO: 2 to SEQ

ID NO: 10, an analogue or a fragment thereof.

Use according to claim 1 or 2, wherein said amino acid sequence is represented by a sequence selected from SEQ ID NO: 11 to SEQ ID NO: 20, an analogue or fragment thereof, and Preferably, one of SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 20, an analog or a fragment thereof is selected.

Use according to any one of the preceding claims, wherein a decrease in the activity, expression or maturation of said biomarker is indicative of aged skin and / or cutaneous signs of aging, associated with ) (s) or not to a dry skin.

5. Use (i) of at least one amino acid sequence selected from SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, an analogue having a sequence identity of at least 90% with said sequence and a biological activity of the same kind, or a fragment of which comprises from 3 to 48 amino acids of said sequence and which has a biological activity of the same kind, or (ii) at least one nucleic acid sequence as defined in claims 1 or 2, for screening active agents or physical treatments capable of modulating the activity, expression or maturation of said amino acid sequence or said nucleic acid sequence.

6. Use (i) of at least one amino acid sequence as defined in claims 1 to 3, or (ii) at least one nucleic acid sequence as defined in claims 1 or 2, for screening active agents or physical treatments for the aged skincare and / or the prevention and / or treatment of cutaneous signs of aging, associated or not with cutaneous dryness, capable of modulating the activity, the expression or the maturation of said skin sequence. amino acid or nucleic acid sequence.

7. Use (i) of at least one amino acid sequence as defined in claims 1 to 3, or (ii) at least one nucleic acid sequence as defined in claims 1 or 2, for characterizing the effectiveness of a cosmetic treatment of the skin.

8. Use according to the preceding claim, wherein the cosmetic treatment is a cosmetic treatment of the aged skin and / or cutaneous signs of aging, associated (s) or not with a dry skin.

9. Cosmetic use of an effective amount (i) of at least one amino acid sequence as defined in claims 1 to 3, or (ii) at least one nucleic acid sequence as defined in claims 1 or 2, as an active agent for preventing and / or treating aged skin and / or cutaneous signs of aging, associated or not with a dry skin.

10. Use according to any one of claims 1 to 4 and 8 and 9, wherein the cutaneous signs of aging are selected from thinning of the skin, loss of firmness, loss of elasticity, loss of density, or loss of skin tone, dryness of the skin, marked microrelief of skin, formation and / or presence of fine lines and / or wrinkles, alteration of radiance of complexion skin, papery appearance of the skin, discolouration of the skin, sagging skin, and wilting of the skin.

11. Use according to any one of claims 1 to 4 and 8 to 10, wherein the signs of cutaneous dryness are chosen from a skin withered, a lack of elasticity, flexibility and / or tone of the skin, a rough touch, crevices, peeling, scaling, wrinkles and fine lines associated with dry skin, and tightness and / or itching.

12. Use of an effective amount of (i) at least one amino acid sequence as defined in claims 1 to 3, or (ii) at least one nucleic acid sequence as defined in claims 1 or 2, or at least one agent modulating the activity, expression or maturation of said amino acid sequence or of said nucleic acid sequence for preparing a pluristratified epithelial cell model, preferably a reconstructed skin model.

13. In vitro or ex vivo method for characterizing a state of the aged skin and / or the cutaneous signs of aging, associated (s) or not with a dry skin comprising at least the steps of:

a) performing, in an isolated sample of a skin, a qualitative or quantitative measurement of the expression, the maturation or the activity of said amino acid sequence as defined in claims 1 to 3 or of said sequence nucleic acid as defined in claims 1 or 2, and

b) comparing said measurement made in step a) with a reference measurement.

The use of any one of claims 1 to 4 and 8 to 10, or the method of claim 13, wherein the aged skin is selected from chronically aged skins and / or photoinduced aging. .

15. A method of in vitro or ex vivo screening of active agents or physical treatments capable of modulating the activity, expression or maturation of an amino acid sequence chosen from SEQ ID NO: 12, SEQ ID NO : 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, an analog having an identity sequence of at least 90% with said sequence and a biological activity of the same kind, or a fragment thereof comprising from 3 to 48 amino acids of said sequence and having a biological activity of the same nature, or a sequence nucleic acid as defined in claims 1 or 2, comprising at least the steps of:

c) performing a qualitative or quantitative measurement of the expression, the maturation or the activity of said amino acid sequence or said nucleic acid sequence, and (d) compare that measure with a reference measure.

16. Process for in vitro or ex vivo screening of active agents or physical treatments for the care of the aged skin and / or the prevention and / or treatment of cutaneous signs of aging, associated or not with an cutaneous dryness, capable of modulating the activity, expression or maturation of an amino acid sequence as defined in claims 1 to 3, or a nucleic acid sequence as defined in claims 1 or 2 , comprising at least the steps of:

(d) compare that measure with a reference measure.

17. A cosmetic process for preventing and / or treating aged skin and / or cutaneous signs of aging, associated or not with cutaneous dryness in an individual in need, comprising at least one step of administering said individual, at least one composition comprising as active agent (i) at least one amino acid sequence as defined in claims 1 to 3, or (ii) at least one nucleic acid sequence as defined in claims 1 or 2.

18. A cosmetic process, in vitro or ex vivo, for characterizing the effectiveness of a cosmetic skin treatment in an individual in need, comprising at least the steps of:

a) performing, prior to the implementation of the cosmetic treatment, in a first isolated skin sample taken from said individual, at least a first qualitative or quantitative measure of the expression, the maturation or the activity of at least an amino acid sequence as defined in claims 1 to 3, or at least one nucleic acid sequence as defined in claims 1 or 2, b) performing, after the implementation of the cosmetic treatment, in a second isolated skin sample taken from said individual, at least a second measurement, qualitative or quantitative, of the expression, the maturation or the activity of said sequence amino acid or nucleic acid sequence, and

19. An isolated peptide represented by an amino acid sequence selected from SEQ ID NO: 18, an analogue or a fragment thereof.

20. Cosmetic composition comprising a peptide as defined in claim 19 or a nucleic acid sequence encoding such a peptide.